Summary/Abstract Crimean-Congo hemorrhagic fever (CCHF) virus is a ssRNA (-) Nairovirus that produces fever, prostration, and severe hemorrhages in humans. Fatality rates for CCHF range from 5-70% based on phylogenetic variation of the virus, transmission route, and different treatment facilities. Originally identified in Russia and the Congo, CCHF has rapidly spread across large sections of Europe, Asia, and Africa. Recently, U.S. citizen and military traffic has increased substantially to the regions affected by CCHF, specifically South Central Asia. As a result, there is a substantial risk for transmission of CCHF and/or its tick vector to the United States of America. This risk has been recently highlighted by a CCHF death of an American Soldier based in Afghanistan in 209. Currently, there is no FDA approved vaccine or therapeutic for treatment of CCHF. Recent reports have identified a viral homologue of the ovarian tumor protease (vOTU) and implicated its involvement down-regulation of the Interferon type 1 immune response through cleavage of post- translational modifying proteins ubiquitin (Ub) and Ub-like interferon-simulated gene 15 (ISG15). Additionally, homologues of CCHFV's vOTU have been suggested to perform similar roles in the economically devastating ssRNA (+) Arteriviruses, Porcine Respiratory and Reproduction Syndrome and Equine Arteritis viruses. Even plant viruses such as the damaging rice stripe virus have been shown to possess a viral ovarian tumor domain protease homologue. This proposal will implement a cost effective fluorescent-based enzymatic assay to identify molecule probes for a vOTU from CCHFV as well as assess whether the molecular probe scaffold can serve as a basis to selectively inhibit other viral and eukaryotic ovarian tumor domain proteases. Specifically a high-throughput screening campaign using a fluorogenic peptide and the MLPCN library will occur at a MLPCN facility. Molecular probes initially identified for the vOTU from CCHFV will be subsequently optimized using secondary, orthogonal, and tertiary in vitro assays as well as in vivo ones. The resulting information provided by molecular probes inhibiting the function of vOTU from CCHFV will allow the necessary insight into not only the role of CCHF's vOTU, but vOTUs in general, in viral evasion of the innate immune system. Additionally, information gathered from vOTU molecular probes would also assist in determining the practicality of developing prophylactics targeting vOTUs and their eukaryotic superfamily relatives that negatively regulate the human innate response.